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Abstract

Background: Ceramide (Cer) participates in neuronal and oligodendroglial cell death and neuroinflammation. We have previously shown that Cer levels increase in the cerebrospinal fluid (CSF)of patients with subarachnoid hemorrhage (SAH). The goal of this study is to further characterize the origin of Cer after SAH.

Methods: Cer and dihydroceramide (DHC) levels as well as the activity of Acid (ASMase) and Neutral sphingomyelinase (NSMase) were measured in the CSF of SAH patients and controls. The de novo synthesis of Cer was estimated by correlating Cer with DHC levels using Spearman correlation. The activity of intracellular enzymes that regulate Cer homeostasis including ASMase, NSMase, sphingomyelinase synthase (SMS) and glucosylceramide synthase (GCS) was investigated using the rat model of SAH. Brain homogenates were used for in vitro enzyme activity measurement. The results were compared using non-parametric analysis.

Results: A total of 26 SAH patients and 8 controls were included in the study. Compared to controls, SAH patients had higher Cer (12.4±8.8 vs. 53.8±48pmol/ml; p=0.001) and DHC (1.4±1.1 vs. 4.7±3.4pmol/ml; p=0.001) in the CSF. In the SAH group, Cer correlated well with DHC (rho=0.91; p<0.001) suggesting a substantial contribution of the de novo pathway to Cer production. ASMase activity in the CSF of human subjects was higher in SAH than in controls (15.04±4.75 vs.9.62±3.88 IF/μl.min; p=0.017) indicating an increased turnover of sphingomyeline into Cer. Additionally, compared to the controls (n=4), brain samples from SAH animals (n=5) had higher ASMase (15.7±2.1 vs. 9.55±1.53 IF/μg.min; p=0.025) and GCS (1.38±0.35% vs. 0.70±0.04%; p=0.025) enzyme activity. On the other hand, NSMase (1.78±0.55 vs. 1.02±0.75 IF/μg.min; p>0.05) and SMS (7.1±0.9% vs. 9.1±3.8%; p>0.05) activities were similar in both groups.

Conclusions: The increase of Cer after SAH may be explained by the elevation in both ASMase activity and the de novo Cer biosynthetic pathway. GCS transforms Cer into less toxic glycosphingolipids, suggesting that this enzyme may constitute an endogenous Cer detoxifying mechanism.